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PubMed1995

High-performance liquid chromatographic-electrospray mass spectrometric determination of morphine and its 3- and 6-glucuronides: application to pharmacokinetic studies.

A rapid and selective assay of morphine and its 3- and 6-glucuronides in serum, based on high-performance liquid chromatography-electrospray mass spectrometry has been developed. The analytes and the internal standard, codeine or naltrexone, were subjected to solid-phase extraction, using ethyl solid-phase extraction columns, prior to chromatography. A reversed-phase column and a gradient mobile phase consisting of water and methanol were used. The mass spectrometer was operated in the selected-ion monitoring mode. The following ions were used: m/z 286 for morphine, m/z 300 for codeine, m/z 342 for naltrexone, and m/z 462 for morphine 3- and 6-glucuronides. The limit of quantitation observed with this method was 10 ng/ml morphine, 50 ng/ml morphine-6-glucuronide and 100 ng/ml morphine-3-glucuronide. The present method proved useful for the determination of serum levels of the parent drug and its metabolites in pain patients, heroin addicts and in morphine-treated mice.

Animals↗
PubMed1995

Simultaneous quantification of opiates, cocaine and cannabinoids in hair.

The present paper describes a sensitive method developed in our laboratory for the simultaneous analysis of opiates (morphine, codeine and monoacetylmorphine), cocainis (cocaine and benzoylecgonine) and cannabinoids (delta 9-tetrahydrocannabinol and 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid) in hair samples. After decontaminating the sample with dichloromethane, two consecutive hydrolyses were performed in order to achieve the best conditions for extracting the three kinds of drugs from the protein matrix. First the opiate and cocainic compounds were extracted by means of a soft acidic hydrolysis with 0.1 N HCl at 50 degrees C overnight and organic solvent extraction at pH 9.2. The cannabinoids need a stronger basic hydrolysis with 11.8 N KOH for 10 min at laboratory temperature. After adding maleic acid, the cannabinoids were extracted with an organic solvent. The derivatization was carried out with heptafluorobutyric anhydride and hexafluoropropanol. Calibration curves were linear between 0.5-100 ng/mg of hair. Recovery and reproducibility were assured. The quantification limits ranged between 0.04-0.26 ng/mg of hair. Seventy hair samples from known drug abusers were cut into 1-cm segments and analyzed by this method. The ranges of measured concentrations (ng/mg) were 0.31-89 for cocaine, 0.1-5.76 for benzoylecgonine, 0.34-45.79 for morphine, 0.45-39.59 for codeine, 0.09-48.18 for monoacetylmorphine, 0.06-7.63 for THC and 0.06-3.87 for THC-COOH. The results of sectional analyses agreed with the self reported drug histories. The usefulness of this method is in assessing earlier drug consumption, and also at the same time obtaining a chronological profile of the consumption of these three types of drugs.

Cannabinoids↗
PubMed1994

Trypanocidal effect of boldine and related alkaloids upon several strains of Trypanosoma cruzi.

The alkaloids boldine, glaucine, predicentrine, apomorphine, coclaurine, norarmepavine and codeine were tested against the epimastigotes of the Tulahuén and LQ strains and the DM 28c clone of Trypanosoma cruzi. The micromolar concentration to inhibit 50% of the culture growth (Tulahuén strain) for apomorphine, glaucine, predicentrine, boldine, norarmepavine, coclaurine and codeine were 29, 90, 85, 110, 310, 580 and > 1000 respectively. Similar values were obtained with the LQ strain and the DM 28c clone. The most active compounds in inhibiting culture growth also inhibited cell respiration, suggesting that these drugs may act by blocking mitochondrial electron transport. The trypanocidal effects of these alkaloids appear to be correlated with their antioxidative activities.

Alkaloids↗
PubMed2005

Modeling drug detection and diagnosis with the 'drug evaluation and classification program'.

In this study, we propose formal models and algorithms to detect drug impairment and identify the impairing drug type, on the basis of data obtained by a Drug Evaluation and Classification (DEC) investigation. The DEC program relies on measurements of vital signs and observable signs and symptoms. A formal model, based on data collected by police officers trained to detect and identify drug impairments, yielded sensitivity levels greater than 60% and specificity levels greater than 90% for impairments caused by cannabis, alprazolam, and amphetamine. For codeine, with a specificity of nearly 90% the sensitivity was only 20%. Using logistic regression, the formal model was much more accurate than the trained officers in identifying impairments from cannabis, alprazolam, and amphetamine. Both the formal model and the officers were quite poor in identifying codeine impairment. In conclusion, the joint application of the DECP procedures with the formal model is useful for drug detection and identification.

Accidents, Traffic↗
PubMed2006

The effect of opiates on the activity of human placental aromatase/CYP19.

Aromatase, cytochrome P450 19, is a key enzyme in the biosynthesis of estrogens by the human placenta. It is also the major placental enzyme that metabolizes the opiates L-acetylmethadol (LAAM), methadone, and buprenorphine (BUP). Methadone and BUP are used in treatment of the opiate addict and are competitive inhibitors of testosterone conversion to estradiol (E(2)) and 16alpha-hydroxytestosterone (16-OHT) to estriol (E(3)) by aromatase. The aim of this investigation is to determine the effect of 20 opiates, which can be administered to pregnant patients for therapeutic indications or abused, on E(2) and E(3) formation by placental aromatase. Data obtained indicated that the opiates increased, inhibited, or had no effect on aromatase activity. Their effect on E(3) formation was more pronounced than that on E(2) due to the lower affinity of 16-OHT than testosterone to aromatase. The K(i) values for the opiates that inhibited E(3) formation were sufentanil, 7 +/- 1 microM; LAAM, 13 +/- 8 microM; fentanyl, 25 +/- 5 microM; oxycodone, 92 +/- 22 microM; codeine, 218 +/- 69 microM; (+)-pentazocine, 225 +/- 73 microM. The agonists morphine, heroin, hydromorphone, oxymorphone, hydrocodone, propoxyphene, meperidine, levorphanol, dextrorphan, and (-)-pentazocine and the antagonists naloxone and naltrexone caused an increase in E(3) formation by 124-160% of control but had no effect on E(2) formation. Moreover, oxycodone and codeine did not inhibit E(2) formation and the IC(50) values for fentanyl, sufentanil, and (+)-pentazocine were >1000 microM. It is unlikely that the acute administration of the opiates that inhibit estrogen formation would affect maternal and/or neonatal outcome. However, the effects of abusing any of them during the entire pregnancy are unclear at this time.

Aromatase↗
PubMed2005

Automated multiple development thin-layer chromatography for separation of opiate alkaloids and derivatives.

There are three types of opiate alkaloids. First, the poppy alkaloids: morphine, codeine, thebaine, noscapine and papaverine; then, the semi-synthetic and synthetic derivatives used in therapy as antitussives and analgesics, such as pholcodine, ethylmorphine and dextromethorphan; at last narcotic compounds, diacetylmorphine (heroin) and opiates employed as substitutes in treatment of addiction: buprenorphine and methadone. For classical thin-layer chromatography (TLC) of opium alkaloids, it is necessary to use complex eluents with strong alkaline substances to obtain a clean separation between morphinan and isoquinoline compounds. This study purposes the planar chromatographic analysis of these substances by the automated multiple development (AMD) compared with results obtained by classical TLC method. The aim of this work was to achieve the best separation of these opiate alkaloids and derivatives by this modern technique of planar chromatography. The AMD system provided a clean separation for each of three opiates groups studied and the best results have been obtained with universal gradient: methanol 100, methanol-dichloromethane 50/50, dichloromethane 100, dichloromethane 100, hexane 100 for opium alkaloids and with gradient A: 5% of 28% ammonia in methanol 100, acetone 100, acetone 100, ethyl acetate-dichloromethane 50/50, dichloromethane 100 for antitussives and substitutes. Two reagents were used for the detection of alkaloids by spraying: Dragendorff and iodoplatinate reagents. The detection limits with these two reagents were 1 microg for ethylmorphine, thebaine, papaverine, codeine, and 2 microg for morphine and noscapine and other alkaloids.

Antitussive Agents↗
PubMed2005

Surfactant enhanced liquid-phase microextraction of basic drugs of abuse in hair combined with high performance liquid chromatography.

The aim of this study was to evaluate the performance of a technique for simultaneous testing of hydrophilic abuse drugs in hair. The analysis of, codeine and methadone in morphine hair included incubation in methanol (5h, 50 degrees C), Surfactant enhanced liquid-phase microextraction (SE-LPME) and HPLC analysis. This study has demonstrated that SE-LPME constitute a real alternative to the other liquid-phase microextraction methods, for pre-concentration and extraction of hydrophilic drugs in biological samples and has shown the advantages of these optimized methodologies over the traditional microextraction techniques. For these drugs recoveries in the range of 57.5-93.7 were obtained from hair. The drugs were enriched by a factor of 61-128 during SE-LPME. Linearity (r2, 0.9982-0.9997) was obtained in the range of 50-500 microg/l for morphine and 10-500 microg/l for codeine and methadone.

Chromatography, High Pressure Liquid↗
PubMed2005

Oral fluid collection: the neglected variable in oral fluid testing.

The potential to use oral fluid as a drug-testing specimen has been the subject of considerable scientific interest. The ease with which specimens can be collected and the potential for oral fluid (OF) drug concentrations to reflect blood-drug concentrations make it a potentially valuable specimen in clinical as well as forensic settings. However, the possible effects of the OF collection process on drug detection and quantification has often been over looked. Several studies have documented that drug-contamination of the oral cavity may skew oral fluid/blood drug ratios and confound interpretation when drugs are smoked, insufflated or ingested orally. OF pH is predicted to have an effect on the concentration of drugs in OF. However, in a controlled clinical study, the effect of pH was less than that of collection technique. Mean codeine OF concentrations in specimens collected a non-stimulating control method were 3.6 times higher than those in OF collected after acidic stimulation. Mean codeine concentrations were 50% lower than control using mechanical stimulation and 77% of control using commercial collection devices. Several factors should be considered if a commercial OF collection device is used. In vitro collection experiments demonstrated that the mean collection volume varied between devices from 0.82 to 1.86 mL. The percentage of the collected volume that could be recovered from the device varied from 18% to 83%. In vitro experiments demonstrated considerable variation in the recovery of amphetamines (16-59%), opiates (33-50%), cocaine and benzoylecgonine (61-97%), carboxy-THC (0-53%) and PCP (9-56%). Less variation in collection volume, volume recovered and drug recovery was observed intra-device. The THC stability was evaluated in a common commercial collection protocol. Samples in the collection buffer were relatively stable for 6 weeks when stored frozen. However, stability was marginal under refrigerated conditions and poor at room temperature. Very little has been published on the efficacy of using IgG concentration, or any other endogenous marker, as a measure of OF specimen validity. Preliminary rinsing experiments with moderate (50 mL and 2 x 50 mL) volumes of water did not reduce the OF IgG concentration below proposed specimen validity criteria. In summary, obvious and more subtle variables in the OF collection may have pronounced effects on OF-drug concentrations. This has rarely been acknowledged in the literature, but should to be considered in OF drug testing, interpretation of OF-drug results and future research studies.

Drug Stability↗
PubMed2004

Validated assay for the determination of markers of illicit heroin in urine samples for the control of patients in a heroin prescription program.

A fully validated procedure for the simultaneous determination of morphine (MOR), morphine-3-glucuronide (M3G), morphine-6-glucuronide (M6G), 6-acetylmorphine (6AM), codeine (COD), codeine-6-glucuronide (C6G), acetylcodeine (AC), noscapine (NOS) and papaverine (PAP) based on liquid chromatography followed by electrospray mass spectrometry applying multiple reaction monitoring (LC-ESI-MS/MS) in urine samples is described. The extraction was carried out on a Zymark Rapid Trace Workstation using C18 solid-phase extraction cartridges. The separation was performed in 19 min on an Agilent 1100 HPLC system, using a Phenomenex C18 AQUA column (4 microm, 150 mm x 2 mm), which is coupled with an Applied Biosystems API 2000 mass spectrometer. Deuterated analogues were used as internal standards. The limits of detection were in the range of 0.1 ng/ml (PAP) to 7.4 ng/ml (M6G), the coefficients of correlation were higher than 0.996, the precisions ranged from 3% to 12% and the absolute recoveries were between 45% (M3G) and 98% (MOR). Analyses of samples from patients of a heroin prescription program demonstrated the usefulness of the procedure for the analytical differentiation between prescribed synthetic heroin (diamorphine) use and non-prescription heroin abuse on the basis of urine analysis. After the ingestion of pharmaceutical heroin only general markers for heroin use were detected, which are MOR, M3G, M6G and 6AM, respectively. When illicit heroin was abused, additionally to further general markers (COD, C6G) specific markers for non-prescription heroin abuse (AC, NOS, PAP) were found. However, it must be kept in mind that only AC may be regarded as absolute specific marker of non-prescription heroin, because all other compounds may appear in urine after ingestion of opiate alkaloids containing medicines or foods (e.g. poppy seeds). Therefore, patients of a heroin prescription program should be advised not to ingest such products.

Calibration↗
PubMed2006

Patterns of pain and analgesic use in 3- to 7-year-old children after tonsillectomy.

Tonsillectomy is a common surgical procedure usually associated with moderate to severe pain. Although self-report is the gold standard for pain assessment, researchers have not studied young children at home with self-report measures. The purpose of this study was to describe the patterns of self-reported pain intensity and analgesic administration in 3- to 7-year-old children undergoing tonsillectomy during the operative day in the hospital and the first 2 postoperative days at home in Iceland. As part of a larger study, 68 children undergoing tonsillectomy were taught to use the Wong-Baker FACES Pain Rating Scale. Pain intensity scores and data about administration of analgesics were collected from children, the medical record, and the parents over a 3-day period. Children received primarily acetaminophen or acetaminophen with codeine in the hospital. At home, 99% of doses administered were acetaminophen only. Most doses were administered rectally. Forty percent of children received 24-hour therapeutic doses in the hospital. Only 10% received a 24-hour therapeutic dose at home despite significant pain scores of 4 or 5 persisting through the second postoperative day. Younger children were less likely to receive acetaminophen with codeine. In the hospital, children with pain intensity scores of 4 or 5 received prescribed morphine only 13% of the time. Children experienced clinically significant pain through the second postoperative day and will probably require a change in protocol to provide more aggressive pain management earlier. This study extends to younger children the research evidence that current pain protocols are inadequate.

Adult↗
PubMed2001

Pain control in medical abortion.

OBJECTIVES: In patients having medical abortions with methotrexate and misoprostol: (1) to determine if giving ibuprofen or acetaminophen plus codeine with misoprostol (prior to onset of pain) prevented severe pain; and (2) to determine if there were predictors of medical abortion pain. METHODS: A group of 281 women randomized to receive placebo, ibuprofen or acetaminophen with codeine. This was taken at home with the misoprostol 4-6 days after the methotrexate. RESULTS: There were no significant differences between the three groups with respect to age, gestational age, parity, anxiety, depression, worst period pain score, and ethnicity. There was no significant difference with respect to rates of severe pain scores. The mean pain score for the entire group was 6.2 on a scale of 0 to 10. Severe pain (scores of 9 or 10) were reported by 23.4% of women and in this group, the mean maternal age was lower (P=0.05), parity was lower (P=0.01), worst period pain scores were higher (P=0.001), anxiety scores were higher (P=0.05) and satisfaction was lower (P=0.01). CONCLUSIONS: The pain experienced in medical abortion causes significant distress and more research is needed to reduce it.

Abortion, Induced↗
PubMed1992

Evaluation of cutaneous responses and lung function from exposure to opiate compounds among ethical narcotics-manufacturing workers.

We recently demonstrated morphine-6-hemisuccinate-human serum albumin conjugate (M-6-HS-HSA)-specific IgG in serum from ethic narcotics-manufacturing workers. In this article, we present results of epicutaneous tests to opiate compounds and lung-function studies in these same workers. Thirty-nine workers, exposed to opiates, were evaluated for possible work-related changes in lung function and were administered a questionnaire concerning opiate exposure and health history in February 1988. In December 1988, 33 employees with occupational exposure to opiates, six other workers (New Jersey referent) employed at the same factory with minimal exposure to opiate compounds, and 17 nonexposed individuals from Cincinnati, Ohio, were subjected to epicutaneous threshold testing with a panel of six opiate compounds and nine common aeroallergens. In opiate-exposed workers, significantly lower epicutaneous threshold concentrations were detected (compared to New Jersey referent and Cincinnati control subjects) for dihydrocodeine (p less than 0.01), hydrocodone (p less than 0.05), codeine (p less than 0.01), and morphine (p less than 0.05). Significant associates existed among epicutaneous threshold concentrations between the agents tested; that is, individuals with a positive morphine skin test would generally have a positive codeine skin test, etc. Atopic status (positive cutaneous test results to two or more of nine common aeroallergens) was not significantly associated (p greater than 0.05) with positive opiate skin sensitivity. Although the mean cross-shift decrements in FEV1 for all workers were nonsignificant, five opiate-exposed individuals demonstrated cross-shift decrements in FEV1 of greater than 10%. Daily maximum-minus-minimum changes in workweek PEFR (PEFRmax-min) were significantly reduced for Monday through Thursday (p less than 0.05) compared to PEFRmax-min changes during a nonwork, nonexposure 3-day weekend. Ten exposed workers demonstrated daily PEFRmax-min changes of greater than 20%, suggesting acute airway obstruction. Increased cutaneous reactivity to opiate compounds among opiate-exposed workers may reflect development of pharmacologic hyperresponsiveness to opiate compounds.

Adult↗
PubMed1997

Evaluation of the discriminative stimulus and reinforcing effects of dihydroetorphine.

These experiments examined whether dihydroetorphine (DHE) could serve as a reinforcer in rhesus monkeys and evoke the discriminative stimulus effects of heroin (HER) in rats, two procedures useful in predicting the overall abuse potential of compounds. Rhesus monkeys were trained to self-administer i.v. HER (10 micrograms/kg for monkeys M-BA, M-NI, and M-HO; 3 micrograms/kg for monkey M-PO) during daily; 2-h experimental sessions under FR 10 Timeout 4 min schedules. VEH and doses of HER (1-30 micrograms/kg), codeine (COD; 30-1000 micrograms/kg), and DHE (1-100 ng/kg) were then substituted for the HER maintenance doses. Results indicated that DHE served as a reinforcer. The dose of DHE that maintained peak numbers of infusions was 171 and 8571 times smaller than those maintaining peak numbers of infusions of heroin and codeine, respectively. Additionally, male Sprague-Dawley rats were trained to discriminate 0.3 mg/kg HER s.c. from vehicle (VEH) in an FR 10 (fixed-ratio 10), food-reinforced, operant procedure. During tests, HER, morphine (MOR), and DHE dose-dependently evoked heroin-lever responding with ED50s of 0.055, 0.74, and 0.00033 mg/kg, respectively. These results indicate that DHE is self-administered by rhesus monkeys, and potently produces the discriminative stimulus effects of HER in rats, and suggest that DHE would have a substantial potential for abuse.

Analgesics, Opioid↗
PubMed1986

Simultaneous quantification of ethylmorphine O-deethylase and N-demethylase activity by high-performance liquid chromatography.

The N-demethylation and O-deethylation of ethylmorphine by cytochrome P-450 are simultaneously measured using high-performance liquid chromatography. All the metabolites and the substrate are extracted from the enzymatic incubation mixture with isopropanol-methylene chloride (20:80) containing 6.0 micrograms/ml codeine sulfate as an internal standard. Separation of the compounds is achieved on a C18 reversed-phase column using a mobile phase of 1% acetic acid--acetonitrile (85:15) with 1-hexanesulfonic acid as a counter-ion. Total run time is 12 min at a flow-rate of 2.0 ml/min and 144 bar. Assay of ethylmorphine N-demethylase and O-deethylase activities in rat liver microsomes revealed close agreement between this method and conventional ones. N-Demethylation was found to greatly exceed O-deethylation in liver microsomes from either control or phenobarbital-treated rats confirming results from other laboratories. This method can also be used to measure the N- and O-demethylation of codeine.

Animals↗
PubMed1999

Antitussive effect of Adhatoda vasica extract on mechanical or chemical stimulation-induced coughing in animals.

The antitussive activity of Adhatoda vasica (AV) extract was evaluated in anaesthetized guinea pigs and rabbits and in unanaesthetized guinea pigs. AV was shown to have a good antitussive activity. Intravenously, it was 1/20-1/40 as active as codeine on mechanically and electrically induced coughing in rabbits and guinea-pigs. After oral administration to the guinea-pig the antitussive activity of AV was similar to codeine against coughing induced by irritant aerosols.

Animals↗
PubMed2001

Drug contamination of US paper currency.

It is known that US paper currency in the general circulation is contaminated with cocaine. Several mechanisms have been offered to explain this finding, including contamination due to handling during drug deals and the use of rolled up bills for snorting. Drug is then transferred from one contaminated bill to others during counting in financial institutions. The possibility of contamination of currency with other drugs has not been reported. In this study, the author reports the analysis of 10 randomly collected US$ 1 bills from five cities, for cocaine, heroin, 6-acetylmorphine (6-AM), morphine, codeine, methamphetamine, amphetamine and phencyclidine (PCP). Bills were immersed in acetonitrile for 2h prior to extraction and GC-MS analysis. Results showed that 92% of the bills were positive for cocaine with a mean amount of 28.75+/-139.07 microg per bill, a median of 1.37 microg per bill, and a range of 0.01-922.72 microg per bill. Heroin was detected in seven bills in amounts ranging from 0.03 to 168.50 microg per bill: 6-AM and morphine were detected in three bills; methamphetamine and amphetamine in three and one bills, respectively, and PCP was detected in two bills in amounts of 0.78 and 1.87 microg per bill. Codeine was not detected in any of the US$ 1 bills analyzed. This study demonstrated that although paper currency was most often contaminated with cocaine, other drugs of abuse may be detected in bills.

Chromatography, High Pressure Liquid↗
PubMed1997

On cosmetically treated hair--aspects and pitfalls of interpretation.

Popular hair cosmetic treatments like bleaching or permanent waving were found to affect the stability of incorporated drugs and to cause alterations of the fibers at an ultrastructural level. This may result in a partial or complete loss of drug substances, depending on the particular drug molecule and on its concentration prior to the cosmetic treatment. Moreover, from literature, there is some evidence that drug molecules are not only incorporated into the growing fiber by passive diffusion from blood into the matrix cells and melanocytes, but that the substances enter the hair also via perspiration such as sweat and sebum. Since permed and bleached hair shows an enhanced sorption capacity, the risk of false positives or an unusually high drug concentration in cosmetically treated hair was under investigation. Virgin, permed, mildly as well as severely bleached tresses were exposed to artificial sweat or sebum containing cocaine, benzoylecgonine, 6-acetylmorphine, morphine and codeine (500 ng/g). Except codeine, the concentrations measured by GC/MS were very small and quite close to the detection limit indicating a minor importance of drug uptake into hair fiber from the endogenous-exogenous shunt via sebum or sweat. From the results it is concluded that an increased risk of false positive results in hair analysis on bleached and permanent waved hair fibers does exist, but is not particularly severe.

Drug Interactions↗
PubMed2000

Reversed-phase high-performance liquid chromatography for evaluating the distribution of pharmaceutical substances in suppository base-phosphate buffer system.

The aim of this study was to assess the potency of the reversed-phase high-performance liquid chromatography (RP-HPLC) for in vitro evaluation of the distribution behavior of common drugs between one of the generally used suppository bases Witepsol H15 and the rectal liquid which is imitated by a phosphate buffer, pH 7.2. The distribution coefficients (log K) of nine compounds--paracetamol, caffeine, diclofenac, propyphenazone, indomethacin, codeine base, codeine phosphate, phenobarbital acid and phenobarbital sodium salt were determined by the classical shake-flask' method followed by RP-HPLC quantitative assay. The capacity factors log k' of the compounds were determined on reversed-phase C18 column at a number of methanol-5 mM phosphate buffer, pH 7.2 mobile phases containing different percentages of methanol (phiMeOH). The apparent capacity factors log k(w)app were derived by extrapolation of the methanol concentration to zero and using the correction for ionization, the real capacity factors log k'(w) were calculated. The lipophilicity of the compounds was assessed by the partition coefficients CLOGP and the distribution coefficients CLOGD7.2, calculated for the n-octanol-water system. Correlations between log k'(w) and CLOGP, log k(w)app and CLOGD7.2, log k(w)app and log K were found. The last correlation indicated that the parameter log k(w)app was suitable for evaluating the distribution behavior of the studied drugs in the examined Witepsol H15-rectal liquid system. The predictive power of this correlation was tested by a set of nine non-congeners. It was shown that the classical 'shake-flask' method for determination of the distribution behavior of the studied drugs between the suppository base Witepsol H15 and the phosphate buffer, pH 7.2 might be replaced by the RP-HPLC technique due to its priorities of rapid, stable and reproducible experiments.

Buffers↗